Abstract: This study was performed to determine lower osmotic pressure (0.075 M KCl) duration optimal for breaking granulosa cell membrane and to determine its effect on reconstructed embryos formation, reconstructed embryos developmental potential and apoptosis in blastocysts in bovine nuclear transfer by intraplasmic injection. Enucleated bovine oocytes were divided into five groups according to holding time of 0.075 M KCl to granulosa cells: control (0), 30, 60, 90 and 180 sec. There were significant differences in reconstructed embryos formation between different holding time of 0.075 M KCl and control (p<0.05) and control had low rate of reconstructed embryos (27.4% versus 41.7, 36.4, 41.0 and 36.0%). The 180 sec experiment group had significant low rate of cleaved embryos and blastocysts (p<0.05). The 90 and 180 sec groups had significant high rate of apoptotic blastomere and they were 10.4 and 11.6% (p<0.05), respectively. These results suggested that lower osmotic pressure to bovine granulosa cells could accelerate the formation of reconstructed embryos in nuclear transfer by intraplasmic injection and long holding time of lower osmotic pressure had disadvantageous effect on blastocysts through resulting in low blastocysts rate and high level of apoptosis in blastocysts.